Notwithstanding the great progresses in the allopathic medicine, the whole world has always shown great interest in the use of medicinal plants to treat their sicknesses and diseases.
One of the main diseases is diabetes mellitus which affects 5% of the population worldwide. Diabetes mellitus is a disorder in the metabolism of carbohydrates and lipids that has a hereditary character and that contributes to increasing the death rate due to vascular, neurological and renal disorders, among others. This pathology is also associated to infertility in both sexes as well as to erectile dysfunction in men, so its prevention is important to prevent such sequels.
Another main condition in women is post-menopause, in which the sexual and depressive symptoms are the most severe (Amore et al., 2007), observing a decline in memory (Elsabath et al., 2005) and greater fatigue. Another important condition that affects women is osteoporosis, which can lead to impairment and death.
Men are affected by the prostatic hyperplasia, an important condition. The percentage of population affected by this condition is 50% at 50 years of age, increasing to 80% towards the end of the eighth decade of life.
Given the problematic mentioned above, the existence and use of a food supplement that attacks each of the conditions mentioned above and that, thanks to its properties, resulted in an energizing, and memory and learning enhancer complement would be ideal.
Peru is characterized by a great vegetal biodiversity, combined with an ancestral culture of its use for medicinal purposes that is very deeply-rooted in the Andean and Amazonian peoples. However, this traditional medicine is mono-herbal; that is, each health problem is treated with one plant, unlike the Hindu, Chinese and Korean cultures where mixtures of several plants (poly-herbal) are frequently used to solve health problems. Another difference is that the traditional Peruvian knowledge is transmitted orally, and little or none of it is recorded in reliable texts. Ignorance of writing in the pre-Hispanic age is one of the causes of the absence of much traditional information, although part of it has been included in written documents as of the Spanish conquest, particularly through the chroniclers.
The representative varieties of Peru used in the present invention are maca (Lepidium meyenii) and yacon (Smallanthus sanchifolius), each of which has interesting therapeutic properties and which will be discussed in detail below.
MACA
Generalities
Maca (Lepidium meyenii) is a plant of the Brassicaceae family originating in the central Andes of Peru, characterized by its capacity to grow in areas above 4000 meters of altitude where other plants hardly grow. Maca has different varieties characterized by the external color, of which 13 varieties have been identified in the area of Carhuamayo, Junin (Peru) ranging from white to black (Tello et al., 1992). Yllescas (1994) has studied the primary components of three varieties: red, yellow and black. Differences are found regarding the content of pure protein, soluble sugars by direct reduction, riboflavin, potassium and iron (Yllescas, 1994;Gonzales, 2006).
It is presumed that the domestication process of maca began two thousand years ago in Ondores, Peru (Matos, 1975; see Gonzales, 2006a). The cultivation of maca is centered in Chincaycocha, a region located in the central Andes of Peru as described in the conquest chronicles and in subsequent reports (Cieza de León, 1553;Cobo, 1653;Ruiz, 1952). By the second half of the 20th century, maca continued being described as exclusive of the central Andes (León, 1964).
The hypocotyls are the edible part of the plant, which comprises the bulbous part that grows under the cotyledons and is similar to a radish of approximately 8 cm diameter and that grows under the ground. In 2003, Marin-Bravo described that the maca grown in the coastal area does not develop the reserving organ that characterizes the hypocotyls obtained in the central Andes.
For its consumption, traditionally, the maca hypocotyls are first dehydrated naturally, its diameter being reduced practically to 3-4 cm, washed with warm water, then soaked in boiled water overnight and then subject to cooking during one or two hours with the same liquid it was soaked in until obtaining a dark brown liquid (Córdova, 2003). Another ancestral custom is to cook the maca in a clay pot during the night, let it settle overnight and finally consume it whole or blended with the same liquid it was boiled in (Córdova, 2003;Gonzales, 2006).
It has been scientifically shown that naturally dried and boiled maca has energizing properties, properties on memory and learning, and also therapeutic properties for osteoporosis, prostatic hyperplasia and in the improvement of fertility. The descriptions about the biological effects of black maca and red maca appear as a result of the experimental work and published as of 2005 (Gonzales et al., 2005; 2006; 2007; 2008;Rubio et al., 2007).
Extraction Processes
Chacón (1961) uses 3 alkaloidal extracts to verify the biological effect of maca on fertility. The methodological design is, however, poor and has been criticized by others (León, 1964;Zheng et al., 2000; Gasco et al., 2008) so it is difficult to take into account any association between these extracts and the biological activity.
Zheng et al. (2000) has described an extraction method of a lipid fraction containing 20 to 30% of fatty acids of Lepidium and 10% of macamides (U.S. Pat. No. 6,267,995; Zheng et al., 2000).
Diverse producers use the ethanolic maceration stage as described by Tello and Porras, (1999) or in U.S. Pat. No. 6,267,995.
In the inventors' laboratory, the effect of a synthetic macamide, N-benzyl-5-oxo-6E, 8E-octadecadienamide, has been tested, which had no effect in the count or daily production of spermatozoids (Gonzales, 2006 page 104). Recently in the inventors' laboratory, it has not been observed either that another macamide, N-benzyl-hexadecanamide, has an effect on the number of spermatozoids in mice unlike the atomized hydroalcoholic extract of black maca that significantly increases it. Most of the biological activity is rather found in the more polar fractions (Gonzales, 2006; 2006a; Valerio & Gonzales, 2005).
Zolezzi (1997) mentions that dried and washed maca can be macerated in alcohol. The maceration can be done by introducing 20 g to 40 g of maca per liter of alcohol and letting macerate for at least five days. However, due to the characteristics of maca, this is not advisable, since the proteins, minerals, as well as some carbohydrates are not soluble in such an extract and they would be lost, unless the remaining filtrate is dried and reused. This process will extract the alkaloids, as well as some soluble glycosides. The biological activity of this preparation is not mentioned.
Garró et al., 1993 has achieved separating four alkaloid fractions from dry and pulverized hypocotyls of the plant; however, the alkaloids responsible for the biological effects of the maca have not been identified. It is necessary to specify that maca may have antagonist effects, for example, black maca increases the mitosis or is anti-apopthotic and increases the production of spermatozoids (Gonzales et al., 2006) while red maca decreases the mitosis or is apopthotic (Gonzales et al., 2005).
Chacón (1961) in his Bachelor thesis “Phytochemical Study of Lepidium meyenii Walp” indicates the process of extraction of secondary metabolites from 50 g of the pulverized maca product corresponding to hypocotyls grated and desiccated in the stove at 70° C.-75° C. during 12 hours, which is subjected through a soxhlet to the successive action of the solvents such as acetone, ether, alcohol and distilled water. The tests are carried out at the boiling temperature of the solvent. Chacón (1961) concludes that he has found three alkaloids, starch, glucides, fatty acids, tanines and scarce concentration of saponines. In the conclusions he mentions that the preliminary observations of the administration of the alkaloidal extract of Lepidium meyenii to rats and toads shows the following effects: a) Increase in the procreation of albino rats; b) Clear and marked stimulation of the follicular maturation also in albino rats; c) No effect on the spermatogenesis induced in the toad. The design of Chacón (1961) has been objected by other researchers (León, 1964, Zheng et al., 2000;Ruiz-Luna et al., 2005; Gasco et al., 2008) due to the small number of animals employed (in most cases it refers only to one animal), the time of administration, since the biological markers were subjective more than quantitative, and finally it does not use statistical analyses. Moreover, the effects over the spermatogenesis have been clearly established in studies in the inventors' laboratory (Gasco et al., 2007;Gonzales 2006; 2006a; Rubio et al., 2006a).
Regarding the extraction with water, no specification exists about the optimum water volume and the time of cooking (Gonzales et al., 2006a; Córdova, 2003). Cóndor (1991) prepares extracts base on 100 g of maca in 300 ml of water and finds out that the treatment reduces the number of empty or aborted ewe hoggs. Zolessi (1997) describes maca is boiled in the same amount of water during thirty to sixty minutes. Then it can be blended with the cooking water adding other ingredients or the maca can simply be consumed separately and the cooking water as a beverage. He does not say if this preparation has any verifiable biological activity.
Maca Products
The commercial products of maca include tablets, pills, capsules, flours, liquors, tonics and mayonnaise (Li et al., 2001), which are mostly constituted by maca flour with or without gelatinization. In the case of tablets or capsules, these contain between 450 and 500 mg of maca hypocotyls and the amount prescribed of 3 to 6 tablets or capsules per day would correspond to 3 grams of hypocotyls, a value that is very far from the average daily consumption of an inhabitant of the central Andes, which is estimated in 20 to 40 grams. To consume 40 grams of hypocotyls a person would have to take more than 80 capsules or tablets per day.
In the case of maca flour, whether dry or fresh, since it has not undergone the traditional preparation process it lacks the therapeutic properties or in any case such properties are diminished. On the other hand, such maca flour has a low solubility in water, which is improved in gelatinized maca. The solubility in water is optimum if the product proceeds from aqueous or boiled hydroalcoholic extraction processes.
The maca flour has a strong flavor, very peculiar in this plant, which is not acceptable by many people not used to its use (De Rivero and Ustariz, 1897). The foregoing problem is solved by adding juices and other mixtures to disguise its flavor (Quiroz and Aliaga, 1997). Another inconvenience is that the consumption of maca flour causes digestive discomforts.
Biological Activity
Besides the well-known nutritional effects (Canales et al., 2000), on the spermatogenesis (Gonzales et al., 2001; 2001a) and fertility (Cobo, 1653;Chacón, 1961;Cóndor, 1991), the energizers, antidepressants, anxiolytics (Gonzales, 2006; 2006a), tranquilizers (Tapia et al., 2000; Gonzales 2006; 2006a) and antioxidants of maca (Sandoval et al., 2002), have demonstrated some particular properties in varieties of maca such as the black and the red maca. It has recently been demonstrated that red maca reduces the size of the prostate in the benign prostatic hyperplasia experimentally induced in rodents (Gonzales et al., 2005; 2007) and reverts the osteoporosis in ovariectomized rats (Carla Gonzales, not published, see FIG. 1). On the other hand the black maca increases the number of spermatozoids, see Table 2 in a quantity greater than with other varieties of maca such as the yellow or red maca (Gonzales C. et al., 2006), and has an effect on the improvement of memory and learning in the model of ovariectomized animals, which simulates what occurs in post-menopause women (Rubio et al., 2006), or in the model of animals treated with scopolamine, which experimentally simulates the Alzheimer Disease (Rubio et al., 2007). The black and red varieties of maca have an important antioxidant activity, which represents a potential food supplement advisable for people's health (Gonzales, not published).
Secondary Metabolites
In many cases it has been intended to associate the presence of secondary metabolites such as macaenos and macamides (Zheng et al., 2000), prostaglandins, sterols, and amides of poly-unsaturated fatty acids in the hypocotyls-root (Li et al., 2001) of maca with its biological properties. Thus, the properties of maca to improve fertility have suggested that they could be due to the presence of biologically active isothiocyanates derived from the hydrolysis of glucosinolates specifically due to the benzyl-isothiocyanate and the p-methoxybenzyl isothiocyanate (Johns, 1981;Li et al., 2001). However, there is no study wherein a specific compound of maca is isolated and that administered to experimental animals has demonstrated any biological effect, reason why any statement of the biological properties of such or such compound falls on the speculation ground. Moreover, the metabolites of the glucosinolates have pro-apopthotic and anti-proliferative properties, an effect completely opposed to the required one to increase fertility (Valreio & Gonzales, 2005).
The glucosinolates measurement is currently used to standardize the maca products. Since glucosinolates are metabolized both in the plant and in the gastrointestinal tract into isothiocyanates and these in turn into other metabolites in the organism (Gonzales & Valerio, 2006; Fahey et al., 2001), not being stable compounds they would not be adequate chemical markers. This has been demonstrated in the inventors' laboratory, where different batches with red maca with the same amount of glucosinolates for each batch show different biological responses (Gonzales, not published).
Yacon
Generalities
Yacon (Smallanthus sanchifolius) is an asteraceae from the Andean areas that grows in zone of not more than 3,000 meters of altitude and the culture of which has expanded to other latitudes. In Peru it is found especially in humid temperate areas in Andean slopes, in dry inter-Andean valleys, as well as in the coast. Both the tubercle and the leaves have hypoglycemic properties, properties of improvement of the lipid profile as well as of intestinal absorption of calcium. The tuberous roots of yacon accumulate almost 10%, based on the fresh weight, of inulin type fructooligosaccharides (FOSs), which are known as food ingredients with health benefits (Narai-Kanayama et al., 2007), the main saccharide being beta-1,2-oligofructane (Valentova et al., 2004). The potential of the yacon root to treat hyperglycemia, see tables 3 and 4, renal problems and for skin rejuvenation seem to be due to the oligofructanes, while the antioxidant, anti-hyperglicemic and cytoprotecting activities of the yacon leaves seem to be due to the content of phenols (Valentova and Ulrichova, 2003; Simonovska et al., 2003). The yacon root is also known for its prebiotic capacity (Pedreschi et al., 2003). Another property of yacon is that of oxygenation of the bisphenol A (Yoshida et al., 2002), and endocrine disrupter that is present in plastics, thus producing a decrease or inhibition of its noxious action to the organism.
Low temperature, high humidity storage of yacon is recommended; even so, there is a transformation of the FOSs into fructose (Narai-Kanayama et al., 2007). The tuberous roots of yacon contain phenolic, flavonoid, alkaloid, steroid, glycosides, carbohydrates compounds (Alvarez et al., 2008) and diterpenoids (Dou et al., 2008). It is possible to find the inulin in 7.8% of hydrolyzed extracts and 7.01% in non-hydrolyzed extracts (Alvarez et al., 2008).
Biological Activity
The phenolic acids of yacon seem to be responsible for its effect on the metabolism of glucose. It has been demonstrated that the caffeic, chlorogenic, rosmarinic and ferulic acids reduce the production of glucose acting over the gluconeogenesis and glucogenolysis. It has recently been demonstrated in the asteraceae, from which yacon comes from, that the phenolic compounds and the antioxidant activity increase as the growing altitude increases (Spitaler et al., 2008).
Most of the phenolic compounds studied increase the levels of the glucokinase RNAm similar to how insulin does (Valentova et al., 2007). Five derivatives of the caffeic acid have been detected in the aqueous extract of the yacon root (Takenaka et al., 2003), which may be responsible for the properties observed in this plant. The yacon roots also show an important effect by producing a positive balance of calcium and magnesium, and thus obtaining a greater osseous mineral retention (Lobo et al., 2007). The sub-chronic use for 4 months of yacon root flour in male rats was well tolerated and did not show adverse effects or toxicity at a dose of 340 mg and 6800 mg FOS/bodily weight. Under these conditions the triglyceride levels decrease although no effects were observed over the glycemia (Genta et al., 2005). The hypoglycemic effect of yacon is not due to a lower intestinal absorption of glucose (Matsuura et al., 2004). The yacon root also has antioxidant properties and among the antioxidants are the chlorogenic acid and the tryptophano (Yan et al., 1999).
The leaves have also shown having a favorable effect on the reduction of the blood glucose levels.
Extraction Processes and Biological Activity
Trying to look for better methods to extract the polysaccharides in the yacon leaves, the effect of the extraction with microwaves and with the traditional extraction with boiling water has been proven. A better extraction has been found by using microwaves at a rate of 280 W twice during 15 minutes each time (Li et al., 2007). Sesquiterpenes lactones have been found in the leaves, which inhibit the production of nitric acid induced by lipopolysaccharides (LPS) in murine RAW 264.7 macrophage cells (Hong et al., 2008). These sesquiterpene lactones have also shown antibacterial activity (Lin et al., 2003). Fluctuanine has the greatest antibacterial activity against Bacillus subtilis between six sesquiterpene-lactoses tested (Lin et al., 2003). The boiled aqueous extract of yacon leaves also shows an antioxidant effect of free radicals and an inhibitory activity over the lipid peroxidation in homogenized rat brain (Tereda et al., 2006). Two organic fractions and two fractions of aqueous extracts of yacon leaves show a high protective effect against the oxidative damage to primary cultures of rat hepatocytes, and a reduced hepatic production of glucose through gluconeogenesis and glucogenolisis (Valentova et al., 2004). The total content of phenols in the extracts range from 10.7 to 24.6% (Valentova et al., 2005). Two fractions of yacon leaves extracts with high content of polyphenols have a high antioxidant power (Valentova et al., 2003).
A 10% decoction of yacon produces a significant reduction of the glucose levels in normal rat plasma when the administration is intraperitoneal or in the digestive tract. Similarly, an administration of 10% of the yacon decoction produces a decrease in the levels of glucose during an oral tolerance to glucose. On the other hand, no effect was observed in diabetic rats induced by streptozotocine after the administration of the 10% yacon decoction. However, the administration of 2% yacon tea ad libitum instead of water for 30 days produced a significant hypoglycemic effect in diabetic rats induced by streptozotocine. After 30 days of administration of the tea, the diabetic rats showed improvements in the plasmatic glucose and insulin, in the bodily weight, and in renal parameters such as kidney weight/bodily weight, depuration of creatinine, urinary excretion of albumin, as compared to control diabetic rats (Aybar et al., 2001).